ABSTRACT The precise surgical resection of breast cancer directly influences patient prognosis. However, it is often difficult or impossible for surgeons to distinguish breast tumor from normal tissue during resection without any intraoperative image guidance. Accurate identification of tumor margins and complete removal of the tumor at the initial surgical operation will be of considerable help in a breast tumor operation. Recently, optical imaging is gaining traction in the image-guided surgery field especially when coupled with near infrared (NIR) fluorescence agent. NIR fluorescent image-guided surgery also affords surgeons numerous advantages such as real-time detection with high-resolution image, and flexible instrument without ionizing radiation. Furthermore, NIR imaging with addition of enhanced-contrast visualizes disease prior to transection, and does not change the appearance of the surgical field of view. Indocyanin green (ICG) is a NIR fluorescence agent and currently registered by the FDA for clinical applications, but not for oncology. In order to use non-toxic ICG as an intraoperative tumor marker, development of a tumor- targeting carrier is essential. Many attempts have been made in the development of carrier molecules, in particular, cell-penetrating peptides (CPPs) have shown to be a promising strategy for improving the delivery and intracellular uptake of diagnostic and therapeutic agents. We have identified that a non-toxic CPP, p28, preferentially enters and is retained significantly longer in breast cancer cells. We hypothesize that developing p28 as a nontoxic tumor-targeting carrier for ICG will provide an ideal intraoperative imaging agent to clearly distinguish tumor and surrounding normal tissue. Preliminary data with this new imaging molecule (p28 chemically conjugated to ICG, ICG-p28) support the hypothesis. We will take a strategic approach to test our hypothesis in clinically relevant setting in multiple breast cancer animal models. It potentially provides significant impact based on unique approach to prevent recurrence and consequently reduce risk of metastatic disease and mortality.